The pDERMATT (plasmid DNA encoding recombinant MART-1 and tetanus toxin fragment-c) plasmid is a novel in-house developed anti-cancer vaccine which encodes a melanoma associated epitope (Mart-1) and an immuno stimulatory sequence (tetanus toxin fragment-c). The pharmaceutical development of pDERMATT necessitated the availability of an assay for the quantification and purity determination of pDERMATT active pharmaceutical ingredient (API), the produced bulk drug and its pharmaceutical dosage form. An anion-exchange liquid chromatographic method (AEX-LC) with ultraviolet (UV) detection was developed, which is based on separation on a non porous anion-exchange (NPR AEX) column with a mobile phase gradient of 0.45-0.53M NaCl in 20mM Tris-HCl 10% isopropanol (IPA) pH 9 and UV detection at 260 and 280nm. The method was found to be precise, accurate and linear over a concentration range of 5-150microg/ml. The supercoiled topoisoform of pDERMATT was well separated from the linear and open-circular form, the main degradation products formed during stress testing, confirming its stability-indicating capability. The use of photo diode array (PDA) detection enabled us to confirm all visible peaks to contain DNA. Additionally capillary gel electrophoresis (CGE) showed the same peak profile as the developed HPLC method. The developed LC-UV method will be used for the pharmaceutical quality control of pDERMATT API, bulk drug and its pharmaceutical dosage form.